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JoVE Journal
Biology
단일 세포 다중 오믹스 실험을 위한 마우스 경동맥 내강에서 내피 세포 분리
단일 세포 다중 오믹스 실험을 위한 마우스 경동맥 내강에서 내피 세포 분리
JoVE Journal
Biology
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JoVE Journal Biology
Isolation of Endothelial Cells from the Lumen of Mouse Carotid Arteries for Single-Cell Multi-Omics Experiments

단일 세포 다중 오믹스 실험을 위한 마우스 경동맥 내강에서 내피 세포 분리

Full Text
3,887 Views
09:12 min
October 4, 2021

DOI: 10.3791/63128-v

Sandeep Kumar*1, Aitor Andueza*1, Nicolas Villa-Roel1, Juyoung Kim1, Dong-Won Kang1, Hanjoong Jo1,2

1Wallace H. Coulter Department of Biomedical Engineering,Georgia Institute of Technology and Emory University, 2Division of Cardiology,Georgia Institute of Technology and Emory University

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Overview

This study presents a novel protocol for isolating endothelial cells and nuclei from mouse carotid arteries under various flow conditions, enabling detailed examination of molecular pathways related to atherosclerosis. The method enhances the ability to perform single-cell omics experiments, facilitating the study of endothelial cell responses to disturbed blood flow.

Key Study Components

Research Area

  • Single-cell omics
  • Endothelial cell biology
  • Atherosclerosis research

Background

  • Endothelial cells are difficult to study due to their low abundance in blood vessels.
  • The protocol significantly improves the yield of endothelial enriched samples.
  • Understanding molecular pathways influenced by blood flow dynamics is crucial for therapeutic development.

Methods Used

  • Isolation of endothelial cells and nuclei from carotid arteries of mice.
  • In vivo experimentation with various flow conditions.
  • Utilization of enzymatic digestion and subsequent cell processing techniques.

Main Results

  • Successful extraction and analysis of single endothelial cells for gene expression profiling.
  • Identification of responses to both acute and chronic disturbed flow conditions.
  • Establishment of potential therapeutic targets for atherosclerosis.

Conclusions

  • This method demonstrates a robust approach for studying endothelial cells in a single-cell context.
  • Findings have significant implications for understanding vascular biology and developing treatments for vascular diseases.

Frequently Asked Questions

What is the significance of isolating endothelial cells?
Isolating endothelial cells allows for targeted studies of their functions and responses in various physiological and pathological environments.
How does disturbed flow affect endothelial cells?
Disturbed blood flow can lead to changes in gene expression that contribute to the development of vascular diseases, such as atherosclerosis.
What techniques are used in this study?
The study employs enzymatic digestion, centrifugation, and single-cell sequencing techniques to analyze isolated cells.
Can this method be applied to other vascular tissues?
Yes, the method can potentially be adapted to isolate endothelial cells from other blood vessels.
What are potential applications of this research?
The research can inform the development of new therapies targeting endothelial function in various cardiovascular conditions.
What are some challenges in carotid artery isolation?
Challenges include the meticulous dissection required to avoid damage and contamination during the isolation process.
What is the role of flow conditions in vascular biology?
Flow conditions are crucial in determining endothelial cell behavior, which influences vascular health and disease.

우리는 단일 세포 체학 실험을 수행하기 위해 안정하거나 방해받는 흐름 조건에 노출 된 마우스 경동맥의 내강에서 내피 세포와 핵을 분리하는 방법을 제시합니다.

혈관에서는 내피 세포의 수가 매우 적어 연구하기가 어렵습니다. 우리의 프로토콜은 이러한 한계를 제거하고 새로운 분자 경로와 치료법을 연구 할 수있게합니다. 우리의 프로토콜을 통해 내피가 풍부한 샘플을 얻고 급성 및 만성 교란 흐름이이 세포 유형에 어떤 영향을 미치는지 연구 할 수 있습니다.

내피 세포를보다 자세히 연구 할 수있는 능력은 표적 유전자를 정의 할 수있게하고, 따라서 내피 세포를 표적으로하는 죽상 동맥 경화증에 대한 잠재적 인 치료제를 정의 할 수 있습니다. 경동맥 격리는 까다 롭고 인내와 경험이 필요합니다. 시작하려면 부 프레 노르 핀 킬로그램 당 0.05 밀리그램을 피하에 주사하여 동물을 준비하십시오.

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